Petersen J, Dandri M, Bürkle A, Zhang L, Rogler C E
Marion Bessin Liver Research Center, Department of Medicine, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
J Virol. 1997 Jul;71(7):5455-63. doi: 10.1128/JVI.71.7.5455-5463.1997.
Persistent hepadnavirus infection leads to oxidative stress and DNA damage through increased production of toxic oxygen radicals. In addition, hepadnaviral DNA integrations into chromosomal DNA can promote the process of hepatocarcinogenesis (M. Feitelson, Clin. Microbiol. Rev. 5:275-301, 1992). While previous studies have identified preferred integration sites in hepadnaviral genomes and suggested integration mechanisms (M. A. Buendia, Adv. Cancer Res. 59:167-226, 1992; C. E. Rogler, Curr. Top. Microbiol. Immunol. 168:103-141, 1991; C. Shih et al., J. Virol. 61:3491-3498, 1987), very little is known about the effects of agents which damage chromosomal DNA on the frequency of hepadnaviral DNA integrations. Using a recently developed subcloning approach to detect stable new integrations of duck hepatitis B virus (DHBV) (S. S. Gong, A. D. Jensen, and C. E. Rogler, J. Virol. 70:2000-2007, 1996), we tested the effects of increased chromosomal DNA damage induced by H2O2, or of the disturbance in DNA repair due to the inhibition of poly(ADP-ribose) polymerase (PARP), on the frequency of DHBV DNA integrations. Subclones of LMH-D21-6 cells, which replicate DHBV, were grown in the presence of various H2O2 concentrations and exhibited up to a threefold increase in viral DNA integration frequency in a dose-dependent manner. Moreover, inhibition of PARP, which plays a role in cellular responses to DNA breakage, by 3-aminobenzamide (3-AB) resulted in a sevenfold increase in the total number of new DHBV DNA integrations into host chromosomal DNA. Removal of either H2O2 or 3-AB from the culture medium in a subsequent cycle of subcloning was accompanied by a reversion back towards the original lower frequency of stable DHBV DNA integrations for LMH-D21-6 cells. These data support the hypothesis that DNA damage sites can serve as sites for hepadnaviral DNA integration, and that increasing the number of DNA damage sites dramatically increases viral integration frequency.
持续性嗜肝DNA病毒感染通过增加毒性氧自由基的产生导致氧化应激和DNA损伤。此外，嗜肝DNA病毒DNA整合到染色体DNA中可促进肝癌发生过程（M.费特尔森，《临床微生物学评论》5:275 - 301，1992年）。虽然先前的研究已确定嗜肝DNA病毒基因组中的优先整合位点并提出了整合机制（M.A.比恩迪亚，《癌症研究进展》59:167 - 226，1992年；C.E.罗格勒，《微生物学与免疫学当前主题》168:103 - 141，1991年；C.施等，《病毒学杂志》61:3491 - 3498，1987年），但对于损伤染色体DNA的试剂对嗜肝DNA病毒DNA整合频率的影响知之甚少。我们使用最近开发的亚克隆方法来检测鸭乙型肝炎病毒（DHBV）的稳定新整合（S.S.龚、A.D.詹森和C.E.罗格勒，《病毒学杂志》70:2000 - 2007，1996年），测试了H2O2诱导的染色体DNA损伤增加或聚（ADP - 核糖）聚合酶（PARP）抑制导致的DNA修复干扰对DHBV DNA整合频率的影响。复制DHBV的LMH - D21 - 6细胞亚克隆在不同浓度的H2O2存在下生长，病毒DNA整合频率以剂量依赖方式增加了高达三倍。此外，3 - 氨基苯甲酰胺（3 - AB）对在细胞对DNA断裂反应中起作用的PARP的抑制导致新的DHBV DNA整合到宿主染色体DNA中的总数增加了七倍。在随后的亚克隆循环中，从培养基中去除H2O2或3 - AB伴随着LMH - D21 - 6细胞稳定DHBV DNA整合频率恢复到原来较低水平。这些数据支持以下假设：DNA损伤位点可作为嗜肝DNA病毒DNA整合的位点，并且增加DNA损伤位点的数量会显著提高病毒整合频率。
